Following localized trauma, damaged axons in the adult mammalian central nervous system (CNS) regress, and subsequent regeneration of these damaged axons is very limited. This failure of regeneration strongly impedes recovery of CNS function and contributes to paralysis, sensory dysfunction and cognitive impairment. To date, the study of brain axon regeneration has almost exclusively relied upon postmortem analysis of fixed tissue from intact preparations, which yields static images. These snapshots are suboptimal for evaluating therapeutic interventions, as they often fail to distinguish regenerating axons from sprouting of undamaged fibers or spared axons at the lesion site. We have developed a model system in which long-distance regeneration of axons can be studied with time-lapse imaging in the intact adult mouse brain. Systemic treatment of adult rats with p-chloro-amphetamine (PCA), causes rapid regression of dorsal raphe serotonin axons, followed by a slow return of serotonergic innervation over many weeks. The Linden lab has adapted this PCA protocol to adult BAC transgenic mice in which the complete extent of serotonin neurons is labeled with EGFP. Using a two-photon microscope and a cranial window, we have repeatedly imaged the same volume of neocortex and thereby tracked serotonergic axons before and = 26 weeks after lesion with PCA to provide time-lapse measurements of identified surviving, regressing and regenerating fibers. Here, I propose to extend this model system and shift towards mechanistic inquiry.
Aim 1. Do serotonin axons regenerate following a stab injury to the neocortex? I propose to repeat immunohistochemistry and in vivo time-lapse imaging of serotonin axons, replacing PCA treatment with a glial-scar inducing stab injury. My goal is to have two well-defined model systems for axonal damage and regeneration, one conventional, pan-cellular and glial- scar-forming and the other cell-type-specific and non-scar forming in order to compare molecular interventions and candidate therapies for functional recovery.
Aim 2. Do serotonin neurons of the dorsal raphe have a unique basal gene expression profile that underlies their unusual capacity for axonal regeneration? Or might the crucial gene expression events in these cells only become evident following PCA or stab- evoked injury? I propose to perform single-cell genetic profiling to quantify gene expression patterns within serotonin neurons prior to axonal damage, immediately following PCA or stab-evoked injury, and after axonal regeneration. This will provide candidate genes for manipulation to alter regeneration.
Following stroke or traumatic brain injury in adults, recovery of function is often very limited, resulting in persistent paralysis, sensory dysfunction, cognitiv impairment and disorders of mood. Researchers have sought to develop therapies to promote recovery of function by stimulating the regrowth of axons, the long information-sending extensions of nerve cells in the brain. I propose a project investigating the role of intrinsic and extrinsic factors influencing regenerating axons in the brain of the intact adult mouse, with the specific intent that this research will provide insights into the mechanism of this recovery and potentially reveal genetic targets for therapeutic intervention in axons that fail to regrow.
|Kajstura, Tymoteusz J; Dougherty, Sarah E; Linden, David J (2018) Serotonin axons in the neocortex of the adult female mouse regrow after traumatic brain injury. J Neurosci Res 96:512-526|
|Jin, Yunju; Dougherty, Sarah E; Wood, Kevin et al. (2016) Regrowth of Serotonin Axons in the Adult Mouse Brain Following Injury. Neuron 91:748-762|